Simple and effective strategies for detection of allele dropout in PCR-based diagnosis of Wilson disease.

To the Editor: Drs. Lam and Mak, in a recent article in this journal (1 ), described the mechanisms leading to allele dropout in the PCR-based diagnosis of Wilson disease (WD) and reported potential solutions to this problem. We propose 2 strategies that would enable unequivocal and rapid identification of allele dropout in WD. In WD, an autosomal recessive disorder, mutations in the ATP7B gene lead to accumulation of copper in the liver and the brain (2 ). In PCR-based detection of WD carriers and presymptomatic individuals in affected families, the presence of numerous single-base variations in the ATP7B gene gives rise to allele dropout, the nonamplification of one of the alleles (1 ). In WD compound heterozygotes, if the wild-type allele corresponding to an identified mutation fails to amplify, the PCR product shows apparent homozygosity, leading to faulty diagnosis. Screening the entire ATP7B gene to exclude allele dropout, as recommended by Lam and Mak (1 ), is an arduous job because ATP7B is 80 kb long with 21 exons and a coding length of more than 6.5 kb. The alternative suggestion, to design primers for PCR from intronic sequence-lacking nucleotide variants (1 ), can be hampered by lack of information on single-base variations, because HAPMAP project data (http://www.hapmap.org/) do not provide complete information on all the neutral nucleotide variants in different population groups. We suggest a much simpler and faster method for detection of allele dropout. Because WD is a simple Mendelian disorder that follows an autosomal recessive mode of inheritance, normal parents of a WD pa-